Quantification of total serum transferrin and transferrin sialoforms in human serum; an alternative method for the determination of carbohydrate-deficient transferrin in clinical samples

Carbohydrate deficient transferrin (CDT) is a biochemical marker for congenital disorders of glycosylation (CDG), chronic alcohol consumption, and forensic medicine diagnosis.

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Diagnosis of Congenital Disorders of Glycosylation by Capillary Zone Electrophoresis of Serum Transferrin

Clinical Chemistry ◽

10.1373/clinchem.2003.021568 ◽

2004 ◽

Vol 50 (1) ◽

pp. 101-111 ◽

Cited By ~ 72

Author(s):

Hubert A Carchon ◽

Roland Chevigné ◽

Jean-Bernard Falmagne ◽

Jaak Jaeken

Keyword(s):

Capillary Zone Electrophoresis ◽

Reference Group ◽

Congenital Disorders ◽

Serum Transferrin ◽

Congenital Disorders Of Glycosylation ◽

Zone Electrophoresis ◽

Carbohydrate Deficient Transferrin ◽

Group A ◽

Capillary Zone ◽

Relative Migration

Abstract Background: Congenital disorders of glycosylation (CDG) are usually diagnosed by isoelectric focusing (IEF) of serum transferrin (Tf). The aim of this study was to evaluate capillary zone electrophoresis (CZE) as a diagnostic alternative to IEF. Methods: We performed 792 CZE analyses of Tf, using the CEofixTM-CDT (carbohydrate-deficient transferrin) assay. Peak identification was based on relative migration times (RMTs) to reduce migration variability. Results: Tf profiles comprised three main groups (A–C). Groups A and B were characterized by one or two dominant tetrasialo-Tf peaks, whereas group C showed a widely variable Tf isoform composition. Group A was composed of four subgroups: a major group with a typical Tf profile (considered as reference group), two minor groups with decreased or moderately increased trisialo-Tf isoform, and a group showing the presence of unknown compounds with RMTs similar to mono- and disialo-Tf. However, these compounds were absent on IEF. Group C contained all profiles from patients with confirmed as well as putative CDG. From the reference group, 99% confidence intervals were calculated for the RMTs of the Tf isoforms, and percentiles representing the Tf isoform distributions were defined. Conclusions: All patients with abnormal IEF results and confirmed CDG were identified by CZE; thus, this method can be used as a diagnostic alternative to IEF in a manner suitable for automation. Because whole serum is analyzed, it should be kept in mind that CZE profiles can show substances other than Tf.

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Screening Using Serum Percentage of Carbohydrate-Deficient Transferrin for Congenital Disorders of Glycosylation in Children with Suspected Metabolic Disease

Clinical Chemistry ◽

10.1373/clinchem.2007.093450 ◽

2008 ◽

Vol 54 (1) ◽

pp. 93-100 ◽

Cited By ~ 28

Author(s):

Celia Pérez-Cerdá ◽

Dulce Quelhas ◽

Ana I Vega ◽

Jesús Ecay ◽

Laura Vilarinho ◽

...

Keyword(s):

Metabolic Disease ◽

Pediatric Population ◽

Congenital Disorders ◽

Congenital Disorders Of Glycosylation ◽

Color Development ◽

Combined Use ◽

Carbohydrate Deficient Transferrin ◽

Polymorphic Variants ◽

Glycosylation Defect

Abstract Background: Diagnoses of congenital disorders of glycosylation (CDG) are based on clinical suspicion and analysis of transferrin (Tf) isoforms. Here we present our experience of CDG screening in children with a suspected metabolic disease by determination of serum percentage of carbohydrate-deficient transferrin (%CDT) in tandem with isoelectric focusing (IEF) analysis of Tf and α1-antitrypsin (α1-AT). Methods: We performed approximately 8000 serum %CDT determinations using %CDT turbidimetric immunoassay (TIA). In selected samples, IEF analysis of Tf and α1-AT was carried out on an agarose gel (pH 4–8) using an electrophoresis unit. The isoforms were detected by Western blotting and visualized by color development. We performed neuraminidase digestion of serum to detect polymorphic variants of Tf. Results: We established a cutoff value for serum %CDT of 2.5% in our pediatric population. Sixty-five patients showed consistently high values of serum %CDT. In accordance with Tf and α1-AT IEF profiles, enzyme assays, and mutation analysis, we made the following diagnoses: 23 CDG-Ia, 1 CDG-Ib, and 1 conserved oligomeric Golgi 1 (COG-1) deficiency. In addition, we identified 13 CDG-Ix non Ia, non-Ib; 3 CDG-Ix; and 9 CDG-IIx cases, albeit requiring further characterization; 9 patients with a secondary cause of hypoglycosylation and 6 with a polymorphic Tf variant were also detected. Conclusion: The combined use of CDT immunoassay with IEF of Tf and α1-AT is a useful 1st-line screening tool for identifying CDG patients with an N-glycosylation defect. Additional molecular investigations must of course be carried out to determine the specific genetic disease.

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Nephelometric determination of carbohydrate deficient transferrin

Clinical Chemistry ◽

10.1093/clinchem/42.4.551 ◽

1996 ◽

Vol 42 (4) ◽

pp. 551-557 ◽

Cited By ~ 22

Author(s):

F Schellenberg ◽

M Martin ◽

E Cacès ◽

J Y Bénard ◽

J Weill

Keyword(s):

Sialic Acid ◽

Characteristic Curve ◽

Anion Exchange Chromatography ◽

Receiver Operator Characteristic Curve ◽

Exchange Chromatography ◽

Treatment Center ◽

Serum Transferrin ◽

Receiver Operator Characteristic ◽

Carbohydrate Deficient Transferrin

Abstract We describe a technique for measuring carbohydrate-deficient transferrin (CDT) in serum. Serum transferrin fractions are separated by anion-exchange chromatography on microcolumns. Sialic acid-deficient transferrin fractions are collected in the eluate, and transferrin is then quantified by a rate-nephelometric technique. Imprecision (CV) was 4-5% within-run and 7-9% between runs (n = 15). Comparison with an isoelectric focusing-immunofixation method for transferrin index (x) yielded y = 761x + 7, Sy/x = 39 mg/L. Assay of sera from 90 abstainers or moderate consumers of alcohol showed that 81 (90%) had CDT concentrations between 30 and 70 mg/L. Among 74 alcoholics admitted to an alcohol treatment center, 54 (73%) had CDT > 70 mg/L, i.e., the diagnostic sensitivity was 73% at a specificity of 90% (area under receiver-operator characteristic curve = 0.891).

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Increased Clinical Sensitivity and Specificity of Plasma Protein N-Glycan Profiling for Diagnosing Congenital Disorders of Glycosylation by Use of Flow Injection–Electrospray Ionization–Quadrupole Time-of-Flight Mass Spectrometry

Clinical Chemistry ◽

10.1373/clinchem.2018.296780 ◽

2019 ◽

Vol 65 (5) ◽

pp. 653-663 ◽

Cited By ~ 8

Author(s):

Jie Chen ◽

Xueli Li ◽

Andrew Edmondson ◽

Gail Ditewig Meyers ◽

Kosuke Izumi ◽

...

Keyword(s):

Mass Spectrometry ◽

Electrospray Ionization ◽

Flow Injection ◽

Time Of Flight ◽

Congenital Disorders ◽

Congenital Disorders Of Glycosylation ◽

Detection Range ◽

Flight Mass Spectrometry ◽

Carbohydrate Deficient Transferrin ◽

Glycan Profiling

Abstract BACKGROUND Congenital disorders of glycosylation (CDG) represent 1 of the largest groups of metabolic disorders with >130 subtypes identified to date. The majority of CDG subtypes are disorders of N-linked glycosylation, in which carbohydrate residues, namely, N-glycans, are posttranslationally linked to asparagine molecules in peptides. To improve the diagnostic capability for CDG, we developed and validated a plasma N-glycan assay using flow injection–electrospray ionization–quadrupole time-of-flight mass spectrometry. METHODS After PNGase F digestion of plasma glycoproteins, N-glycans were linked to a quinolone using a transient amine group at the reducing end, isolated by a hydrophilic interaction chromatography column, and then identified by accurate mass and quantified using a stable isotope-labeled glycopeptide as the internal standard. RESULTS This assay differed from other N-glycan profiling methods because it was free of any contamination from circulating free glycans and was semiquantitative. The low end of the detection range tested was at 63 nmol/L for disialo-biantennary N-glycan. The majority of N-glycans in normal plasma had <1% abundance. Abnormal N-glycan profiles from 19 patients with known diagnoses of 11 different CDG subtypes were generated, some of which had previously been reported to have normal N-linked protein glycosylation by carbohydrate-deficient transferrin analysis. CONCLUSIONS The clinical specificity and sensitivity of N-glycan analysis was much improved with this method. Additional CDGs can be diagnosed that would be missed by carbohydrate-deficient transferrin analysis. The assay provides novel biomarkers with diagnostic and potentially therapeutic significance.

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Evolutionary changes in acute-phase proteins in alcoholic hepatocellular diseases.

Clinical Chemistry ◽

10.1093/clinchem/29.1.45 ◽

1983 ◽

Vol 29 (1) ◽

pp. 45-47 ◽

Cited By ~ 4

Author(s):

C Perier ◽

A Chamson ◽

R Engler ◽

J Frey

Keyword(s):

Acute Phase ◽

Acute Phase Proteins ◽

Hepatic Disease ◽

Serum Transferrin ◽

Chronic Alcohol ◽

C Reactive Protein ◽

Chronic Alcohol Consumption ◽

Reactive Protein ◽

Evolutionary Changes ◽

Two Stages

Abstract We studied the pattern of acute-phase proteins (orosomucoid, C-reactive protein, and haptoglobin) in hepatocellular deficiency due to chronic alcohol consumption, characterized by a decrease in serum transferrin concentration. We found that their patterns could vary independently of hepatocellular deficiency, but depend on the progression of hepatic disease. The most useful protein for discriminating the stage of inflammatory reaction is orosomucoid. In moderate hepatocellular deficiency, acute-phase proteins are increased independently of the decrease in transferrin, whereas in severe hepatocellular deficiency the acute-phase proteins are also decreased. Thus, it is possible to distinguish the two stages of hepatocellular deficiency by following changes in the concentration of orosomucoid.

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